Cloning and Analysis of Paternally Expressed Gene EB30 in Zygote of a Tobacco Interspecies Hybrid

doi:10.13560/j.cnki.biotech.bull.1985.2015.01.018

Abstract

Abstract: As known that EB30（EST accession number EB426694）paternally expressed in zygote of a tobacco interspecies hybrid, its CDS sequence was then cloned by using the EST database of NCBI. Also some characters of EB30 protein were analyzed by the methods of bioinformatics. Genome walking and hiTAIR-PCR were used to obtain the 5' flanking region of EB30 including the promoter and 5'UTR. The EB30-EGFP fusion protein expression vector driven by the 5' flanking region of EB30 was constructed and transformed into the tobacco. The results showed that EB30 gene encoded 211 amino-acid residues with a molecular weight 22.616 2 kD and pI5.63. This protein was hydrophilic, and predicted to locate in the mitochondrion. The phylogenetic tree showed that EB30 protein of tobacco was so closed to the tomato and potato, the amino acid homology was about 73%. There were two conserved regions near the N-terminal and C-terminal in homologous proteins from different species. Finally, the 5' flanking region of EB30 was proved to have a promoter activity in the early embryogenesis by fluorescence observation in the zygote and 2-celled proembryo from transgenic plants.

Key words: tobacco, interspecies hybrid, early embryogenesis, paternal expression, cloning

Luo An. Cloning and Analysis of Paternally Expressed Gene EB30 in Zygote of a Tobacco Interspecies Hybrid[J]. Biotechnology Bulletin, 2015, 31(1): 115-121.

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